Luglio Agosto 2017 - Articoli originali

Fabry Disease in Southern Sardinia: epidemiological results from screening in an extensive area


Introduction: Epidemiological data relating to the prevalence and incidence of Fabry disease (FD) and other Lysosomal Storage diseases (LSDs) are largely underestimated and not yet well known. Distribution of the disease varies according to geographical area and to ethnic origin. Heterozygous females are also at risk of contracting severe and multi-symptomatic forms of FD.
Aim: To demonstrate the results obtained in outpatient surgeries situated in an area comprising 319,340 inhabitants.
Methods: Out of a total of 2710 nephrologist visits, 150 patients with suspected FD (73 undergoing dialysis and 77 conservative management) were selected. The relatives of one female patient on dialysis who had tested positive were investigated and a further 11 patients thus identified (total: 4 males and 7 females) within a micro-area of 21,822 inhabitants, i.e. a prevalence rate of one positive case every 1,818 inhabitants. These data relate to the first 18 months of screening.
Conclusions: In the field of nephrology, patients with high levels of proteinuria or microalbuminuria (150-200 mg/day) should be screened for FD, particularly in areas with a high incidence and/or prevalence of kidney disease. Once positive patients of both sexes have been identified, they should immediately be referred for cardiologic and neurological assessment.

Key words: Fabry, familiar heritage, rare disease


Epidemiological data reporting the prevalence and incidence of Fabry disease and other LSDs are largely lacking due to a marked diversity in geographic regions and ethnic populations. Accordingly, as the available data are likely underestimated due to missed diagnoses of these rare disorders, the actual diffusion of FD remains to be clarified [1]. In recent years, following numerous reports of a late onset FD, the main focus has been extended to a series of medical specialities, and to investigating prevalence of the disease in heterozygous individuals. Indeed, a significant number of female carriers may develop Fabry disease–related symptoms. The worldwide prevalence of FD is hard to ascertain, although estimates range from one patient per 40,000 to 60,000 individuals [2], to the one every 200,000 inhabitants reported in a Japanese survey. Calculations however are complicated due to the different phenotypes observed in males and females, including classic type 1, type 2 with later onset and atypical mutations [3]. Fabry disease is an X-linked lysosomal storage disease caused by a deficit of lysosomal enzyme α -galactosidase A (α -Gal A). The majority of males lacking α -Gal A activity develop the classic phenotype of Fabry disease, which affects multiple organ systems. Glycosphingolipids, predominantly globotriaosylceramide (GL-3) and galabiosylceramide, accumulate in the lysosomes of various cells (eg, in the vascular endothelium of multiple organs) owing to α -Gal A deficiency. Accumulation of GL-3 in the lysosomes causes lysosomal and cellular dysfunction, which, in turn, triggers a cascade of cells and tissue ischemia and fibrosis [4]. A recent consensus conference on the KDIGO guidelines to be adhered to in the treatment and prevention of FD has identified the major affected organs and relative symptoms. Currently, the main aim is to implement early treatment, particularly in view of the wide variability of asymptomatic patients and patients affected by severe kidney, heart or neurological conditions [5]. Genetic variants of the disease are becoming increasingly numerous. Indeed, although the majority of patients with Fabry disease are white, the disorder has been described in patients from a series of ethnic groups, including those with Hispanic, African, Asian, and Middle Eastern ancestry [6]. In northern Sardinia, out of a total of 178 individuals with suspected FD, genomic alterations have already been identified in nine patients from two family clusters characterized by severe neurological manifestations (D313Y) and renal and cardiovascular impairment (R227Q) [7]. Routine screening for FD in the general population should be implemented; the test is extremely simple and would greatly benefit FD patients, as early detection of the disease may prevent progressive renal impairment [8]. Reports of isolated family groups affected by FD have also been published [9], thus supporting recommendations that genetic research should be undertaken on even distant relatives of the patient, as suggested by the National Society of Genetic Counsellors [10]. The aim of the present study was to implement routine screening of the patient population attending specialist nephrology clinics.

Materials and methods

In March 2014 our Nephrologic Unit commenced screening for Fabry disease in an extensive area covered by the Cagliari (Italy) Primary Care Trust. The results obtained relate to the first 18 months of screening. The area comprises 319,340 inhabitants and 22 regular Nephrology out-clinics for the treatment and prevention of chronic kidney disease. In the period 2015-2016, approximately 10,803 nephrologist visits were scheduled in both patients who periodically attended the clinic and new patients. Of the original 10,803 patients, 2,712 were identified: 156 patients were subsequently screened, 73 of which on dialysis and 77 outpatients. The main selection criteria for molecular genetic screening were as follows: male and female patients, even in the absence of typical skin alterations [11]; additional criteria included the finding of past or current microalbuminuria at levels of 100- 200 mg/day. All extra-renal symptoms should be carefully investigated, particularly those of a neurological nature [12], including previous transient ischaemic attacks and/or stroke, hypertrophic cardiomyopathy with or without arterial hypertension, arrhythmias, valve disease, acroparesthesia, family history for undetermined kidney disease; patients with type 1 and 2 diabetes should not be excluded from screening.

Exclusion criteria: only patients with a confirmed diagnosis (glomerulonephritis, even when identified by means of electron microscopy, Adult Polycystic Kidney Disease, and other well-known genetic diseases).

The screening program was carried out on whole blood samples as follows: a 1mL blood sample by means of EDTA to test at least 2 μg DNA. Blood samples collected via DBS were extracted for analysis using TripleQuad mass spectrometry. The levels of α-galactosidase were assessed in plasma by spectrofluorometry (normal plasma range A> 2.6 nmol/L; lyso-GB3 concentrations were measured in patients with evocative FD DNA mutation and/or single polymorphism in the GLA gene by Tandem mass-spectrometry (MRM-MS) (CentoFabry, Centogene ®); lyso-GB3 normal range < 1.8 ng/mL [13]. Genetic analysis on α-GAL A or GLA (Centogene ®) was performed by means of high-throughput automated Sanger sequencing enabling a large amount of sequencing data to be generated at the same time. This technique enables genetic analysis of 48 Fabry disease patients in parallel overnight. The Centogene© AG methodology utilizes sequencing platforms from Illumina; before checking GLA full gene sequencing covers the entire coding region, exon/intron boundaries and 200 bp of the gene promoter and after Deletion/duplication analysis/mutational scanning of GLA.

Results of screening

Negative findings were reported for all outpatients and dialysis patients who underwent genetic testing, with the exception of an apparently asymptomatic 65-year old haemodialysis patient with a previous history of no longer identifiable proteinuric kidney disease. The patient tested positive at enzyme and genetic assay. As the patient belonged to a very large family, the screening program was extended to all her known relatives, resulting in the detection of an additional 11 carriers of Fabry disease (4 hemizygous males and 7 heterozygous females) (Figure 1). The mean age of the 12 screened patients was: 37.0 + 21.2 years (1-65 years); mean age of the four males 26.2 + 20.2 years (9-54); and mean age of the eight females 42.4 + 20.8 (1- 65). Figure 2 illustrates the family pedigree.

Epidemiological findings and frequencey

Throughout the extensive screening area comprising 319,340 inhabitants, the above family cluster was identified from the much smaller coastal-mountainous region of Sarrabus-Gerrei, accounting for 21,822 inhabitants (Figure 3).

Based on the initial extended screening area the rate of prevalence was 1 case per 26,612 inhabitants; however, when related to the more confined area of the Sarrabus-Gerrei region, prevalence was calculated as 1 case per 1,818 inhabitants.

This area was already renowned for its high prevalence of terminal chronic kidney disease compared to the rest of the island and the rest of Italy: respectively, Sarrabus-Gerrei: 872 cases per one million inhabitants, Sardinia 810, Italy 580 (2012 Census of the Sardinian Section of the Italian Society of Nephrology).

A unique genetic mutation was detected in all these subjects: c.159C>A (p.Asn53Lys). This mutation has not been described previously in literature either as an atypical mutation, or in view of the correlated symptoms of Fabry disease.


Results of blood chemistry tests:

Alpha-galactosidase levels determined in all 12 patients corresponded to 0.62 + 0.38 (min-max: < 0.2 – 1.2 µmol/L); in males 0.30 + 0.10 ( min-max< 0.2 – 0.4 µmol/L); in females 0.78 + 0.36 ( min-max: 0.2 – 1.2 µmol/L). The 0.2 µmol/L α-galactosidase levels refer to the female patient undergoing haemodialysis – this sample was collected pre-dialysis during the longer interdialytic interval to avoid dialysis-correlated interferences.

LysoGb3 levels in all patients corresponded to 1.6 + 0.6 ng/mL (max-min: 3.2 – 0.9), in females 0.78 + 0.36 ng/mL (max-min: 1.6 – 1.4) and in males 1.9 + 1.04 ng/mL (max-min: 3.2 – 0.9). Table 1 illustrates the entire case history with a brief description of renal and some extrarenal symptoms and enzymatic levels. Study continuation: patients underwent thorough kidney screening and were subsequently referred for cardiologic, neurological and ophthalmic assessment. The female patient (n. 9) will need to be re-assessed to examine the possibility of an incorrect diagnosis of multiple sclerosis and/or a correlation between the two diseases. An estimated rate of prevalence for Fabry disease in the area concerned corresponds to approximately one affected subject per 1,818 inhabitants.



Fabry disease is viewed as an obsolete, although still largely underestimated, disease, and is rarely taken into consideration as a causal factor in illnesses of unconfirmed origin in the fields of nephrology, neurology and ophthalmology. In nephrology clinics greater attention should be focused on microalbuminuria; the finding of levels below 200 mg/day may be the sole indication of the presence of renal FD [14].

In the present case, the finding of a family cluster in a geographically isolated and scarcely populated micro-area with a high prevalence of uremia, stemmed from the results of genetic testing of a patient undergoing dialysis. Linthorst et al. [15] estimated the prevalence of Fabry disease in patients on dialysis at 0.22%. Consequently, screening for Fabry disease in large renal dialysis clinics has been suggested. In France, 106 patients on haemodialysis were screened for α-galactosidase activity: one patient with Fabry disease was identified and a further seven family members of this index case carried the same mutation [8]. A study in Japan reported that 1.2% of males receiving renal dialysis were affected by Fabry disease [16]. Conversely, a recent study by Saito et al. reported a 0.02% prevalence of FD in 5,408 male patients and 3,139 female dialysis patients (0.04% in males, 0.0% in females) [17], thus confirming the wide variability of phenotypic penetrance of the disease. Hereditary transmission of FD in the geographically isolated Sarrabus-Gerrei region in Sardinia is likely due to the marriage to healthy carriers of the disease or to people with compromised organs and/or apparatus dating back previous generations. Indeed, up until one hundred years ago this region was even more isolated and cultural tradition encouraged marriages between close family members. Age at onset and at diagnosis of FD is characterised by a wide variability. Mean ages at onset of symptoms and diagnosis of FD were 9 and 23 years (males) and 13 and 32 years (females), respectively, indicating misleading diagnosis or delays in both sexes [18]. In these patients FD was detected during investigation of concurrent involvement of other major organ systems. Closer monitoring and vigilance may contribute towards reducing or postponing progression of the disease and the need for dialysis or transplantation in patients with FD. Delayed onset forms of FD with initial cardiologic manifestations beyond the age of 40 have been reported [19]. Likewise, delayed onset forms of renal FD have been observed; Wilcox et al. reported alterations in renal function in heterozygous females with a mean age of 46 years [20].

In view of the presence in our family cluster of asymptomatic and/or scarcely symptomatic patients, as well as a patient on dialysis, it has not been possible to establish the clinical outcome and phenotypic expression of the newly detected mutation c.159C>A p.Asn53Lys. Ongoing neurological, cardiologic and ophthalmic studies may help to clarify this issue, although an additional practical problem is represented by the poor collaboration of patients and parents of minors. As FD has produced no discernible symptoms in these subjects, they are rather reluctant, with some (see patient n. 10 in Table 1) even refusing to undergo diagnostic tests or commence replacement enzymatic therapy when needed. Other studies have also reported a lack of symptoms, which may explain diagnostic and therapeutic delays, at times resulting in irreversible kidney damage [21, 22]. Missed diagnoses often occur in childhood, as many of these cases may be clinically silent [23]. The present study also included three children, thus underlining the need to raise awareness amongst paediatricians and parents, with the latter proving particularly reluctant to subject their currently asymptomatic children to a series of diagnostic tests.



Numerous reports published in the literature tend to suggest the absence of a significantly overt correlation between screened enzymatic levels and total accumulation; nevertheless, Consultants in a range of fields should continue to screen all individuals, particularly females, in whom a diagnosis of FD may be suspected [24]. In view of the wide range of classic and atypical variants of FD worldwide, estimation of reliable rates of incidence and prevalence of the disease is virtually impossible. The prevalence of FD may at times be characterized by a somewhat random form of distribution, or affect a series of family groups, thus giving rise to the hypothesis of a genetic recycling between distant relatives in geographically and culturally isolated regions. The imprint is that the mutation c.159C>A (p.Asn53Lys) could have a phenotypic mild and/or belated negative effects on renal function. The currently available innovative screening tools are rapid and easy to use and should be adopted in routine diagnostics in Nephrology. A correct use of these tools will facilitate investigations in subjects attending nephrology clinics, particularly in areas characterized by a high incidence of chronic kidney disease.

All procedures performed in studies involving human participants were in accordance with the ethical standards of the institutional and/or national research committee and with the 1964 Helsinki declaration and its later amendments or comparable ethical standards. Informed consent was obtained from all individual participants included in the study.


We are grateful to Genzyme® and Shire® for their kind contributions. 


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